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Gαq sensitizes TRPM8 to inhibition by PI(4,5)P2 depletion upon receptor activation

Luyu Liu, Yevgen Yudin, Chifei Kang, Natalia Shirokova, View ORCID ProfileTibor Rohacs
doi: https://doi.org/10.1101/410878
Luyu Liu
Department of Pharmacology, Physiology and Neuroscience, Rutgers New Jersey Medical School, Newark NJ 07103
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Yevgen Yudin
Department of Pharmacology, Physiology and Neuroscience, Rutgers New Jersey Medical School, Newark NJ 07103
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Chifei Kang
Department of Pharmacology, Physiology and Neuroscience, Rutgers New Jersey Medical School, Newark NJ 07103
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Natalia Shirokova
Department of Pharmacology, Physiology and Neuroscience, Rutgers New Jersey Medical School, Newark NJ 07103
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Tibor Rohacs
Department of Pharmacology, Physiology and Neuroscience, Rutgers New Jersey Medical School, Newark NJ 07103
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ABSTRACT

Activation of G-protein coupled receptors (GPCRs) was proposed to inhibit the cold and menthol sensitive Transient Receptor Potential Melastatin 8 (TRPM8) channels via direct binding of Gαq to the channel. It is well documented that TRPM8 requires the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2 or PIP2] for activity. It was claimed however that a decrease in cellular levels of this lipid does not contribute to channel inhibition upon receptor activation. Here we show that supplementing the whole cell patch pipette with PI(4,5)P2 reduced inhibition of TRPM8 by activation of Gαq-coupled receptors in mouse dorsal root ganglion (DRG) neurons. Activation of the same receptors induced Phospholipase C (PLC) activation and decreased plasma membrane PI(4,5)P2 levels in these neurons. PI(4,5)P2 also reduced inhibition of TRPM8 by activation of heterologously expressed Gαq-coupled muscarinic M1 receptors. Co-expression of a constitutively active Gαq protein that does not couple to PLC inhibited TRPM8 activity, and in cells expressing this protein decreasing PI(4,5)P2 levels using a voltage sensitive 5’-phosphatase induced a stronger inhibition of TRPM8 activity than in control cells. Our data indicate that PI(4,5)P2 depletion plays an important role in TRPM8 inhibition upon GPCR activation, and Gαq inhibits the channel by reducing its apparent affinity for PI(4,5)P2 and thus sensitizes the channel to inhibition by decreasing PI(4,5)P2 levels.

ACKNOWLEDGEMENTS:

T.R. was supported by NIH grants NS055159 and GM093290. The authors thank Dr. Joshua Berlin for his insightful comments, Dr. David Julius (UCSF) for providing the TRPM8 clone, Dr. David McKemy (University of Southern California) for providing the GFP-TRPM8 mouse line, Dr. Yasushi Okamura (Osaka University, Japan) for providing the ci-VSP and dr-VSP clones, Drs. Nikita Gamper (University of Leeds) and Andrew Tinker (University College London) for providing the tubby-R332H-YFP clone, Dr. Xuming Zhang (Aston University, Birmingham, UK) for providing the 3Gqiq clone, and Linda Zabelka for maintaining the mouse colony.

Footnotes

  • The authors declare no financial conflict of interest

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted September 06, 2018.
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Gαq sensitizes TRPM8 to inhibition by PI(4,5)P2 depletion upon receptor activation
Luyu Liu, Yevgen Yudin, Chifei Kang, Natalia Shirokova, Tibor Rohacs
bioRxiv 410878; doi: https://doi.org/10.1101/410878
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Gαq sensitizes TRPM8 to inhibition by PI(4,5)P2 depletion upon receptor activation
Luyu Liu, Yevgen Yudin, Chifei Kang, Natalia Shirokova, Tibor Rohacs
bioRxiv 410878; doi: https://doi.org/10.1101/410878

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